Subgrading of G2 Pancreatic Neuroendocrine Tumors as 2A (Ki67 3% to < 10%) Versus 2B (10% to ≤ 20%) Identifies Behaviorally Distinct Subsets in Keeping with the Evolving Management Protocols.

BACKGROUND: Grade 1/2 PanNETs are mostly managed similarly, typically without any adjunct treatment with the belief that their overall metastasis rate is low. In oncology literature, Ki67-index of 10% is increasingly being used as the cutoff in stratifying patients to different protocols, although there are no systematic pathology-based studies supporting this approach. METHODS: Ki67-index was correlated with clinicopathologic parameters in 190 resected PanNETs. A validation cohort (n = 145) was separately analyzed. RESULTS: In initial cohort, maximally selected rank statistics method revealed 12% to be the discriminatory cutoff (close to 10% rule of thumb). G2b cases had liver/distant metastasis rate of almost threefold higher than that of G2a and showed significantly higher frequency of all histopathologic signs of aggressiveness (tumor size, perineural/vascular invasion, infiltrative growth pattern, lymph node metastasis). In validation cohort, these figures were as striking. When all cases were analyzed together, compared with G1, the G2b category had nine times higher liver/distant metastasis rate (6.1 vs. 58.5%; p < 0.001) and three times higher lymph node metastasis rate (20.5 vs. 65.1%; p < 0.001). CONCLUSIONS: G2b PanNETs act very similar to G3, supporting management protocols that regard them as potential therapy candidates. Concerning local management, metastatic behavior in G2b cases indicate they may not be as amenable for conservative approaches, such as watchful waiting or enucleation. This substaging should be considered into diagnostic guidelines, and clinical trials need to be devised to determine the more appropriate management protocols for G2b (10% to ≤ 20%) group, which shows liver/distant metastasis in more than half of the cases, which at minimum warrants closer follow-up.

Dataset for the reporting of carcinoma of the exocrine pancreas: recommendations from the International Collaboration on Cancer Reporting (ICCR).

AIMS: Current guidelines for pathology reporting on pancreatic cancer differ in certain aspects, resulting in divergent reporting practices and a lack of comparability of data. Here, we report on a new international dataset for pathology reporting on resection specimens with cancer of the exocrine pancreas (ductal adenocarcinoma and acinar cell carcinoma). The dataset was produced under the auspices of the International Collaboration on Cancer Reporting (ICCR), which is a global alliance of major (inter)national pathology and cancer organisations. METHODS AND RESULTS: According to the ICCR's rigorous process for dataset development, an international expert panel consisting of pancreatic pathologists, a pancreatic surgeon and an oncologist produced a set of core and non-core data items based on a critical review and discussion of current evidence. Commentary was provided for each data item to explain the rationale for selecting it as a core or non-core element and its clinical relevance, and to highlight potential areas of disagreement or lack of evidence, in which case a consensus position was formulated. Following international public consultation, the document was finalised and ratified, and the dataset, which includes a synoptic reporting guide, was published on the ICCR website. CONCLUSIONS: This first international dataset for cancer of the exocrine pancreas is intended to promote high-quality, standardised pathology reporting. Its widespread adoption will improve the consistency of reporting, facilitate multidisciplinary communication, and enhance the comparability of data, all of which will help to improve the management of pancreatic cancer patients.

Cytologic predictors of malignancy in bile duct brushings: a multi-reviewer analysis of 60 cases.

Diagnosing malignancy in bile duct brushings is highly challenging. Seven reviewers of variable backgrounds and levels of participation in bile duct brushing sign out blindly reviewed 60 specimens (30 malignant with histologic confirmation and 30 benign (15 stented) with resection or ≥18 months of uneventful follow-up), testing the utility of 14 malignant characteristics. Eleven characteristics were statistically significantly associated with malignancy including 3-dimensional clusters (63% in malignant vs 3% in benign, odds ratio 50, P=0.0003), pleomorphism (62 vs 3, odds ratio 48, P=0.0004), 2-cell population (60% vs 3, odds ratio 44, P=0.0005), chromatin pattern (hypo/hyperchromasia) changes (70% vs 7%, odds ratio 33, P<0.0001), high nuclear-to-cytoplasmic ratio (48 vs 3%, odds ratio 27, P=0.0023), cytoplasmic vacuoles (43 vs 3%, odds ratio 22, P=0.0042), nuclear irregularity (70 vs 10%, odds ratio 21, P<0.0001), cellular discohesion (38 vs 3%, odds ratio 18, P=0.0082), hypercellularity (23% vs 0), nuclear molding (20% vs 0) and prominent nucleoli (21% vs 0). Necrosis and infiltrating inflammation were not helpful in identifying malignancy ('neutrophil cannibalism' was noted in 43% malignant); 21/30 (70%) malignant brushings had ≥3 malignant characteristics, while 23 (77%) benign brushings had none. Of 20 brushings with ≥4 characteristics, 1(5%) proved benign and showed detachment atypia, a close malignant mimicker in brushings. Identification of 3 characteristics maximized the combined sensitivity (70%), specificity (97%) and accuracy (83%), but sensitivity dropped as number of characteristics increased. Identification of 3/11 characteristics (3-dimensional clusters, pleomorphism, high nuclear-to-cytoplasmic ratio, nuclear irregularity, hypercellularity, discohesion, chromatin changes, vacuoles, prominent nucleoli, molding and 2-cell population) improves pathologists' overall performance greatly.

Non-ampullary-duodenal carcinomas: clinicopathologic analysis of 47 cases and comparison with ampullary and pancreatic adenocarcinomas.

Literature on non-ampullary-duodenal carcinomas is limited. We analyzed 47 resected non-ampullary-duodenal carcinomas. Histologically, 78% were tubular-type adenocarcinomas mostly gastro-pancreatobiliary type and only 19% pure intestinal. Immunohistochemistry (n=38) revealed commonness of 'gastro-pancreatobiliary markers' (CK7 55, MUC1 50, MUC5AC 50, and MUC6 34%), whereas 'intestinal markers' were relatively less common (MUC2 36, CK20 42, and CDX2 44%). Squamous and mucinous differentiation were rare (in five each); previously, unrecognized adenocarcinoma patterns were noted (three microcystic/vacuolated, two cribriform, one of comedo-like, oncocytic papillary, and goblet-cell-carcinoid-like). An adenoma component common in ampullary-duodenal cancers was noted in only about a third. Most had plaque-like or ulcerating growth. Mismatch repair protein alterations were detected in 13% (all with plaque-like growth and pushing-border infiltration). When compared with ampullary (n=355) and pancreatic ductal (n=227) carcinomas, non-ampullary-duodenal carcinomas had intermediary pathologic features with mean invasive size of 2.9 cm (vs 1.9, and 3.3) and 59% nodal metastasis (vs 45, and 77%). Its survival (3-, 5-year rates of 57 and 57%) was similar to that of ampullary-duodenal carcinomas (59 and 52%; P=0.78), but was significantly better than the ampullary ductal (41 and 29%, P<0.001) and pancreatic (28 and 18%, P<0.001) carcinomas. In conclusion, non-ampullary-duodenal carcinomas are more histologically heterogeneous than previously appreciated. Their morphologic versatility (commonly showing gastro-pancreatobiliary lineage and hitherto unrecognized patterns), frequent plaque-like growth minus an adenoma component, and frequent expression of gastro-pancreatobiliary markers suggest that many non-ampullary-duodenal carcinomas may arise from Brunner glands or gastric metaplasia or heterotopic pancreatobiliary epithelium. The clinical behavior of non-ampullary-duodenal carcinoma is closer to that of ampullary-duodenal subset of ampullary carcinomas, but is significantly better than that of ampullary ductal and pancreatic cancers. The frequency of mismatch repair protein alterations suggest that routine testing should be considered, especially in the non-ampullary-duodenal carcinomas with plaque-like growth and pushing-border infiltration.

Massive gastric juvenile-type polyposis: a clinicopathological analysis of 22 cases.

AIMS: Massive gastric polyposis is a rare entity that is often associated with juvenile polyposis syndrome (JPS). The aim of this study was to evaluate the clinicopathological features of 22 patients with abundant gastric juvenile-type or hyperplastic-like polyps. METHODS AND RESULTS: The study included 12 males and 10 females with a median age of 48 years (range: 13-79 years). Fourteen (64%) patients carried a diagnosis of JPS, and three had prior gastrointestinal adenocarcinomas. Patients without known JPS presented at an older median age (60 years versus 40 years; P = 0.0068). Clinical symptoms included nausea, vomiting, and abdominal pain; 23% of patients were asymptomatic. Eighteen cases showed complete or near-complete carpeting of the gastric mucosa by innumerable polyps, ranging from a few millimetres to ~100 mm. Most polyps formed long, bulbous projections and had characteristic histological features, including a smooth outer contour, prominent stromal oedema, and widely spaced, often cystically dilated glands lined by foveolar epithelium; some polyps had less stroma and more hyperplastic foveolar epithelium. All had normal underlying or adjacent mucosa. Four (18%) cases harboured adenocarcinoma, and seven (32%) others showed dysplasia. SMAD4 immunohistochemical staining showed patchy loss in polyps from 19 of 20 cases tested. Five of six (84%) patients tested had a germline SMAD4 mutation. CONCLUSIONS: Massive gastric juvenile-type polyposis can occur in patients with and without known JPS, and may mimic different conditions, such as other polyposis syndromes and Ménétrier disease. Pathologists play an important role in disease classification, as some patients lack a family or personal history of JPS, have few if any colonic polyps, and may not harbour diagnostic germline mutations.

Adenosquamous Carcinoma of the Esophagus: An NCDB-Based Investigation on Comparative Features and Overall Survival in a Rare Tumor.

OBJECTIVES: Esophageal adenosquamous carcinoma (ASC) is a rare tumor with characteristics of adenocarcinoma (AC) and squamous cell carcinoma (SCC), the two most common esophageal cancers. Its behavior is aggressive but poorly understood. Using the National Cancer Database (NCDB), the clinical features and overall survival of ASC were compared with AC and SCC. METHODS: The NCDB was queried for patients with esophageal ASC, AC, and SCC. Univariate association of histology with patient characteristics and overall survival were analyzed and socioeconomic characteristics were balanced. RESULTS: Clinical M stage was 0 in a significantly lower proportion of ASC (69.0%) than in AC (70.9%) or SCC (75.6%) (p < 0.001). Median survival was lower in patients with ASC (9.6 months) than with AC (13.5) or SCC (9.7) and 2-year OS was lower in patients with ASC (23.8%) than with AC (34.6%) or SCC (26.5%) (p < 0.001). The OS hazard ratio for ASC was 1.14 when compared to AC (95% CI = 1.016-1.267, p = 0.025) and 1.10 when compared to SCC, but the latter was not significant (95% CI = 0.980-1.222, p = 0.111). CONCLUSION: ASC is a rare tumor among esophageal carcinomas with a greater burden of metastatic disease than AC or SCC and worse OS than AC.

The prognostic significance of BAP1, NF2, and CDKN2A in malignant peritoneal mesothelioma.

Cytoreductive surgery and hyperthermic intraperitoneal chemoperfusion for patients with malignant peritoneal mesothelioma has resulted in improved disease control and increased survival. Despite these results, there are significant perioperative risks associated with this aggressive procedure that necessitate consideration of prognostic markers during patient selection. The molecular pathogenesis of peritoneal mesothelioma remains relatively unknown, but extrapolation of findings from their pleural counterpart would suggest frequent alterations in CDKN2A, NF2, and BAP1. Homozygous deletions in CDKN2A portend a worse overall survival in peritoneal mesothelioma. However, the prevalence and prognostic significance of NF2 and BAP1 abnormalities has not been studied. Dual-color fluorescence in situ hybridization using CDKN2A and NF2 locus-specific probes and BAP1 immunohistochemistry identified homozygous CDKN2A deletions (n=25, 29%), hemizygous NF2 loss (n=30, 35%), and/or loss of BAP1 protein expression (n=49, 57%) in 68 of 86 (79%) peritoneal mesotheliomas. Homozygous CDKN2A deletions or hemizygous NF2 loss correlated with shorter progression-free survival (P<0.02) and poor overall survival (P<0.03). Moreover, the significance of these findings was cumulative. Patients harboring both homozygous CDKN2A deletions and hemizygous NF2 loss had a 2-year progression-free survival rate of 9% with a median of 6 months (P<0.01) and overall survival rate of 18% with a median of 8 months (P<0.01). By multivariate analysis, combined homozygous CDKN2A deletions and hemizygous NF2 loss was a negative prognostic factor for both progression-free survival and overall survival, independent of patient age, peritoneal cancer index, completeness of cytoreduction, and extent of invasion. In contrast, loss of BAP1 was not associated with clinical outcome. In summary, homozygous deletions in CDKN2A and hemizygous loss of NF2 as detected by fluorescence in situ hybridization would confer a poor clinical outcome and may guide future treatment decisions for patients with peritoneal mesothelioma.

Ampullary carcinoma is often of mixed or hybrid histologic type: an analysis of reproducibility and clinical relevance of classification as pancreatobiliary versus intestinal in 232 cases.

Histologic classification of ampullary carcinomas as intestinal versus pancreatobiliary is rapidly becoming a part of management algorithms, with immunohistochemical classification schemes also being devised using this classification scheme as their basis. However, data on the reproducibility and prognostic relevance of this classification system are limited. In this study, five observers independently evaluated 232 resected ampullary carcinomas with invasive component >3 mm. Overall interobserver agreement was 'fair' (κ 0.39; P<0.001) with complete agreement in 23%. Using agreement by 3/5 observers as 'consensus' 40% of cases were classified as 'mixed' pancreatobiliary and intestinal. When observers were asked to provide a final diagnosis based on the predominant pattern in cases initially classified as mixed, there was 'moderate' agreement (κ 0.44; P<0.0001) with 5/5 agreeing in 35%. Cases classified as pancreatobiliary by consensus (including those with pure-pancreatobiliary or mixed-predominantly pancreatobiliary features) had shorter overall (median 41 months) and 5-year survival (38%) than those classified as pure-intestinal/mixed-predominantly intestinal (80 months and 57%, respectively; P=0.026); however, on multivariate analysis this was not independent of established prognostic parameters. Interestingly, when compared with 476 cases of pancreatic ductal adenocarcinomas, the pancreatobiliary-type ampullary carcinomas had better survival (16 versus 41 months, P<0.001), even when matched by size and node status. In conclusion, presumably because of the various cell types comprising the region, ampullary carcinomas frequently show mixed phenotypes and intratumoral heterogeneity, which should be considered when devising management protocols. Caution is especially warranted when applying this histologic classification to biopsies and tissue microarrays. While ampullary carcinomas with more pancreatobiliary morphology have a worse prognosis than intestinal ones this does not appear to be an independent prognostic factor. However, pancreatobiliary-type ampullary carcinomas have a much better prognosis than their pancreatic counterparts.

Intrapancreatic distal common bile duct carcinoma: Analysis, staging considerations, and comparison with pancreatic ductal and ampullary adenocarcinomas.

Distal common bile duct carcinoma is a poorly characterized entity for reasons such as variable terminology and difficulty in determining site of origin of intrapancreatic lesions. We compared clinicopathologic features of pancreatobiliary-type adenocarcinomas within the pancreas, but arising from the distal common bile duct, with those of pancreatic and ampullary origin. Upon careful review of 1017 pancreatoduodenectomy specimens with primary adenocarcinoma, 52 (5%) qualified as intrapancreatic distal common bile duct carcinoma. Five associated with an intraductal papillary neoplasm were excluded; the remaining 47 were compared to 109 pancreatic ductal adenocarcinomas and 133 ampullary carcinomas. Distal common bile duct carcinoma patients had a younger median age (58 years) than pancreatic ductal adenocarcinoma patients (65 years) and ampullary carcinoma patients (68 years). Distal common bile duct carcinoma was intermediate between pancreatic ductal adenocarcinoma and ampullary carcinoma with regard to tumor size and rates of node metastases and margin positivity. Median survival was better than for pancreatic ductal adenocarcinoma (P=0.0010) but worse than for ampullary carcinoma (P=0.0006). Distal common bile duct carcinoma often formed an even band around the common bile duct and commonly showed intraglandular neutrophil-rich debris and a small tubular pattern. Poor prognostic indicators included node metastasis (P=0.0010), lymphovascular invasion (P=0.0299), and margin positivity (P=0.0069). Categorizing the tumors based on size also had prognostic relevance (P=0.0096), unlike categorization based on anatomic structures invaded. Primary distal common bile duct carcinoma is seen in younger patients than pancreatic ductal adenocarcinoma or ampullary carcinoma. Its prognosis is significantly better than pancreatic ductal adenocarcinoma and worse than ampullary carcinoma, at least partly because of differences in clinical presentation. Use of size-based criteria for staging appears to improve its prognostic relevance. Invasive pancreatobiliary-type distal common bile duct carcinomas are uncommon in the West and have substantial clinicopathologic differences from carcinomas arising from the pancreas and ampulla.

Adenocarcinoma ex-goblet cell carcinoid (appendiceal-type crypt cell adenocarcinoma) is a morphologically distinct entity with highly aggressive behavior and frequent association with peritoneal/intra-abdominal dissemination: an analysis of 77 cases.

High-grade versions of appendiceal goblet cell carcinoids ('adenocarcinoma ex-goblet cell carcinoids') are poorly characterized. We herein document 77 examples. Tumors occurred predominantly in females (74%), mean age 55 years (29-84), most with disseminated abdominal (77% peritoneal, 58% gynecologic tract involvement) and stage IV (65%) disease. Many presented to gynecologic oncologists, and nine had a working diagnosis of ovarian carcinoma. Metastases to liver (n=3) and lung (n=1) were uncommon and none arose in adenomatous lesions. Tumors had various histologic patterns, in variable combinations, most of which were fairly specific, making them recognizable as appendiceal in origin, even at metastatic sites: I: Ordinary goblet cell carcinoid/crypt pattern (rounded, non-luminal acini with well-oriented goblet cells), in variable amounts in all cases. II: Poorly cohesive goblet cell pattern (diffusely infiltrative cords/single files of signet ring-like/goblet cells). III: Poorly cohesive non-mucinous cell (diffuse-infiltrative growth of non-mucinous cells). IV: Microglandular (rosette-like glandular) pattern without goblet cells. V: Mixed 'other' carcinoma foci (including ordinary intestinal/mucinous). VI: goblet cell carcinoid pattern with high-grade morphology (marked nuclear atypia). VII: Solid sheet-like pattern punctuated by goblet cells/microglandular units. Ordinary nested/trabecular ('carcinoid pattern') was very uncommon. In total, 33(52%) died of disease, with median overall survival 38 months and 5-year survival 32%. On multivariate analysis perineural invasion and younger age (<55) were independently associated with worse outcome while lymph-vascular invasion, stage, and nodal status trended toward, but failed to reach, statistical significance. Worse behavior in younger patients combined with female predilection and ovarian-affinity raise the possibility of hormone-assisted tumor progression. In conclusion, 'adenocarcinoma ex-goblet cell carcinoid' is an appendix-specific, high-grade malignant neoplasm with distinctive morphology that is recognizable at metastatic sites and recapitulates crypt cells (appendiceal crypt cell adenocarcinoma). Unlike intestinal-type adenocarcinoma, it occurs predominantly in women, is disguised as gynecologic malignancy, and spreads along peritoneal surfaces with only rare hematogenous metastasis. It appears to be significantly more aggressive than appendiceal mucinous neoplasms.

Prognostic significance of morphological growth patterns and mitotic index of epithelioid malignant peritoneal mesothelioma.

AIMS: The prognostic significance of histological subtyping of epithelioid pleural mesotheliomas has been recently reported, but similar data are lacking for peritoneal mesotheliomas. The aim of this study was to investigate possible relationships between histological growth patterns of epithelioid peritoneal mesotheliomas, clinicopathological features, and patient outcome. METHODS AND RESULTS: Eighty-four cases of chemotherapy-naive epithelioid peritoneal mesothelioma were classified into tubulopapillary, micropapillary, papillary, tubular, solid and trabecular growth patterns. Pathological features such as depth of invasion, lymphocytic host response, mitotic count, nuclear grade, lymphovascular invasion, lymph node metastasis and stromal desmoplasia were analysed. The most common histological patterns were solid (n = 37, 44%), tubulopapillary (n = 24, 29%), and micropapillary (n = 11, 13%). The overall median survival was 36 months. Patients with solid mesothelioma had shorter overall survival (median, 29 months) than patients with tubulopapillary and micropapillary growth patterns (median, 51 and 53 months, respectively; P = 0.053). A high mitotic index (>5 in 50 high-power fields) was found to be associated with poor survival (P < 0.03). A moderate to severe lymphocytic host response was associated with longer median survival (P = 0.13). CONCLUSIONS: Our study highlights the prognostic importance of the solid growth pattern among diffuse epithelioid peritoneal mesotheliomas, and reaffirms mitotic index as a predictor of overall survival.

KRAS mutational analysis and immunohistochemical studies can help distinguish pancreatic metastases from primary lung adenocarcinomas.

Lung metastases from primary pancreatic adenocarcinomas often have mucinous features, which makes them difficult to distinguish from the primary lung adenocarcinoma. We explored the potential utility of KRAS mutational status and immunohistochemical studies in the evaluation of adenocarcinomas in the lungs of patients with known pancreatic cancer. Metastatic pancreatic cancer cases had fewer solitary lung lesions (5 (15%) versus 37 (95%) for lung primaries; P=0.0001), more tumors with pure (100%) mucinous morphology (16 (50%) versus 9 (23%) for lung primaries; P=0.0037), and more frequent KRAS mutations (24 (75%) versus 18 (46%) for lung primaries; P=0.0093). Presence of the KRAS G12C mutation had 96% specificity and positive predictive value for lung adenocarcinoma, whereas G12R was 99% specific for pancreatic cancer with a positive predictive value of 86%. Of the 18 KRAS mutated mucinous lung tumors, only 3 (16%) occurred in nonsmokers. Conversely, of the 19 KRAS mutated pancreatic cancer metastases, 11 (58%) occurred in nonsmokers. The median overall survival was significantly shorter for patients with metastatic tumors when compared with patients with primary mucinous tumors (19 months, 95% confidence interval, 10-28 months versus 55 months, 95% confidence interval, 39-70 months, P=0.005). CK20 and CDX2 positivity supported metastatic pancreatic cancer, whereas TTF-1 positivity supported primary lung adenocarcinoma. In summary, KRAS G12C mutations, TTF-1, and napsin A were associated with primary lung adenocarcinoma, whereas KRAS G12R mutations, CK20, and CDX2 favored pancreatic adenocarcinoma. We showed survival differences for patients whose pancreatic metastases were synchronous versus metachronous to their primary tumors, and for patients with mucinous pancreatic cancer metastases versus primary mucinous lung adenocarcinomas. Differences in KRAS mutations reflect differences in exposure to tobacco smoking and highlight biological differences between two KRAS oncogene-driven cancers.

Loss of SMAD4 staining in pre-operative cell blocks is associated with distant metastases following pancreaticoduodenectomy with venous resection for pancreatic cancer.

BACKGROUND AND OBJECTIVES: Venous resection of locally advanced pancreatic cancer is associated with increased morbidity and mortality; therefore identification of patients most likely to benefit from this aggressive surgical approach is an important goal. Loss of SMAD4 staining on resected specimens has been associated with outcomes. Few studies have evaluated the prognostic significance of SMAD4 staining of pre-operative cell blocks, which would be useful in clinical decision making for patients with locally advanced disease. METHODS: Clinical data were retrospectively evaluated from all patients undergoing pancreaticoduodenectomy with venous resection. Immunohistochemical staining for SMAD4 was performed on pre-operative cell blocks and subsequent post-operative resections. RESULTS: One hundred seventeen patients underwent pancreaticoduodenectomy with venous resection. Sixty had sufficient specimens available for SMAD4 staining. SMAD4 loss was observed in 70% of resections and was associated with earlier time to metastatic disease. Pre-operative SMAD4 loss correlated well with post-operative staining and was associated with six times higher likelihood of developing metastases. CONCLUSION: In this pilot study, preoperative SMAD4 staining showed a strong correlation with postoperative staining and predicted metastases in locally advanced cancer. Preoperative SMAD4 status may be considered as one of several factors when selecting patients most likely to benefit from aggressive en bloc venous resection.

KRAS mutant allele-specific imbalance is associated with worse prognosis in pancreatic cancer and progression to undifferentiated carcinoma of the pancreas.

KRAS codon 12 mutations are present in about 90% of ductal adenocarcinomas and in undifferentiated carcinomas of the pancreas. The role of KRAS copy number changes and resulting KRAS mutant allele-specific imbalance (MASI) in ductal adenocarcinoma (n=94), and its progression into undifferentiated carcinoma of the pancreas (n=25) was studied by direct sequencing and KRAS fluorescence in situ hybridization (FISH). Semi-quantitative evaluation of sequencing electropherograms showed KRAS MASI (ie, mutant allele peak higher than or equal to the wild-type allele peak) in 22 (18.4%) cases. KRAS FISH (performed on 45 cases) revealed a trend for more frequent KRAS amplification among cases with KRAS MASI (7/20, 35% vs 3/25, 12%, P=0.08). KRAS amplification by FISH was seen only in undifferentiated carcinomas (10/24, 42% vs 0/21 pancreatic ductal adenocarcinoma, 0%, P=0.0007). In 6 of 11 cases with both undifferentiated and well-differentiated components, transition to undifferentiated carcinoma was associated with an increase in KRAS copy number, due to amplification and/or chromosome 12 hyperploidy. Pancreatic carcinomas with KRAS MASI (compared to those without MASI) were predominantly undifferentiated (16/22, 73% vs 9/97, 9%, P<0.001), more likely to present at clinical stage IV (5/22, 23% vs 7/97, 7%, P=0.009), and were associated with shorter overall survival (9 months, 95% confidence interval, 5-13, vs 22 months, 95% confidence interval, 17-27; P=0.015) and shorter disease-free survival (5 months, 95% confidence interval, 2-8 vs 13 months, 95% confidence interval, 10-16; P=0.02). Our findings suggest that in a subset of ductal adenocarcinomas, KRAS MASI correlates with the progression to undifferentiated carcinoma of the pancreas.

Integration of KRAS testing in the diagnosis of pancreatic cystic lesions: a clinical experience of 618 pancreatic cysts.

With improvements in abdominal imaging, detection of incidental pancreatic cysts are becoming increasingly common. Analysis of pancreatic cyst fluid from fine-needle aspiration is particularly important in identifying intraductal papillary mucinous neoplasms (IPMNs) and mucinous cystic neoplasms (MCNs), which have significant implications in clinical intervention and follow-up. Previous controlled studies have shown that KRAS mutations in cyst fluid are highly specific for mucinous differentiation in pancreatic cysts; however, this has not been examined in the clinical setting. Over a 6-year study period, 618 pancreatic cyst fluids obtained by fine-needle aspiration at the time of endoscopic ultrasound were tested for KRAS mutations as part of routine evaluation for a cystic neoplasm. Of the 618 specimens, 603 (98%) from 546 patients were satisfactory for molecular analysis. Patients ranged in age from 17 to 90 years (mean, 63.9 years) and were predominantly female (68%). Pancreatic cysts were relatively evenly distributed throughout the pancreas and ranged in size from 0.6 to 11.0 cm (mean, 2.3 cm). Mutations in KRAS were detected in 232 of 603 (38%) aspirates. Although sufficient for molecular analysis, 320 of 603 (53%) specimens were either less than optimal (38%) or unsatisfactory (15%) for cytopathologic diagnosis. Surgical follow-up information was available for 142 (26%) patients and consisted of 53 KRAS-mutated and 89 KRAS-wild-type cysts. Overall, KRAS mutations had a specificity of 100%, but a sensitivity of 54% for mucinous differentiation. When stratified by cyst type, KRAS had a sensitivity of 67% and 14% for IPMNs and MCNs, respectively. In summary, KRAS mutations were highly specific for mucinous differentiation, but were inadequate in identifying MCNs. Future molecular studies and the combination of other fluid markers are required to improve the detection and classification of pancreatic mucinous neoplasms by endoscopic ultrasound fine-needle aspiration.

Phase II study of induction fixed-dose rate gemcitabine and bevacizumab followed by 30 Gy radiotherapy as preoperative treatment for potentially resectable pancreatic adenocarcinoma.

BACKGROUND: Eighty percent of patients with resected pancreatic ductal carcinoma (PDC) experience treatment failure within 2 years. We hypothesized that preoperative fixed-dose rate (FDR) gemcitabine (GEM) combined with the angiogenesis inhibitor bevacizumab (BEV) and accelerated 30 Gy radiotherapy (RT) would improve outcomes among patients with potentially resectable PDC. METHODS: This phase II trial tested induction FDR GEM (1,500 mg/m(2)) plus BEV (10 mg/kg IV) every 2 weeks for three cycles followed by accelerated RT (30 Gy in 10 fractions) plus BEV directed at gross tumor volume plus a 1-2 cm vascular margin. Subjects underwent laparoscopy and resection after day 85. Therapy was considered effective if the complete pathologic response rate exceeded 10 % and the margin-negative resection rate exceeded 80%. RESULTS: Fifty-nine subjects were enrolled; 29 had potential portal vein involvement. Two grade 4 (3.4%) and 19 grade 3 toxicities (32.8%) occurred. Four subjects manifested radiographic progression, and 10 had undetected carcinomatosis. Forty-three pancreatic resections (73%) were performed, including 19 portal vein resections (44%). Margin-negative outcomes were observed in 38 (88%, 95% confidence interval [CI] 75-96), with one complete pathologic response (2.3%; 95% CI 0.1-12). There were seven (6 grade 3; 1 grade 4) wound complications (13%). Median overall survival for the entire cohort was 16.8 months (95% CI 14.9-21.3) and 19.7 months (95% CI 16.5-28.2) after resection. CONCLUSIONS: Induction therapy with FDR GEM and BEV, followed by accelerated BEV/RT to 30 Gy, was well tolerated. Although both effectiveness criteria were achieved, survival outcomes were equivalent to published regimens.

Outcomes with FOLFIRINOX for borderline resectable and locally unresectable pancreatic cancer.

BACKGROUND: Trials examining FOLFIRINOX in metastatic pancreatic cancer demonstrate higher response rates compared to gemcitabine-based regimens. There is currently limited experience with neoadjuvant FOLFIRINOX in pancreatic cancer. METHODS: Retrospective review of outcomes of patients with borderline resectable or locally unresectable pancreatic cancer who were recommended to undergo neoadjuvant treatment with FOLFIRINOX. RESULTS: FOLFIRINOX was recommended for 25 patients with pancreatic cancer, 13 (52%) unresectable and 12 (48%) borderline resectable. Four patients (16%) refused treatment or were lost to follow-up. Twenty-one patients (84%) were treated with a median of 4.7 cycles. Six patients (29%) required dose reductions secondary to toxicity. Two patients (9%) were unable to tolerate treatment and three patients (14%) had disease progression on treatment. Seven patients (33%) underwent surgical resection following treatment with FOLFIRINOX alone, 2 (10%) of which were initially unresectable. Two patients underwent resection following FOLFIRINOX + stereotactic body radiation therapy (SBRT). The R0 resection rate for patients treated with FOLFIRINOX ± SBRT was 33% (55% borderline resectable, 10% unresectable). A total of five patients (24%) demonstrated a significant pathologic response. CONCLUSIONS: FOLFIRINOX is a biologically active regimen in borderline resectable and locally unresectable pancreatic cancer with encouraging R0 resection and pathologic response rates.

Tissue yield and diagnostic efficacy of fluoroscopic and cholangioscopic techniques to assess indeterminate biliary strictures.

BACKGROUND & AIMS: Evaluation of indeterminate biliary strictures typically involves collection and analysis of tissue or cells. Single-operator, peroral, cholangioscopic techniques have been developed that allow for a biopsy sample to be obtained from a specific area of the visualized stricture. We investigated whether standard fluoroscopic-guided and cholangioscopic-directed (SpyGlass Direct Visualization System; Boston Scientific, Natick, MA) biopsy collection provide adequate tissue for histologic assessment. METHODS: We examined 110 consecutive bile duct specimens collected from 89 patients with indeterminate biliary strictures at a single institution using fluoroscopy or cholangioscopy (from October 2007 to March 2010). Because of the small nature of the intraductal biopsy fragments, special procedures were followed in the pathology laboratory to maximize the amount of tissue for histopathology analysis. RESULTS: Only 4 specimens (3.6%) had insufficient material for a diagnosis. More tissue was obtained from standard fluoroscopic-guided than cholangioscopic-directed biopsies (more biopsy fragments, P = .018; larger total biopsy size, P = .001). Fluoroscopy-guided biopsies assessed indeterminate biliary strictures with 76% sensitivity and 88% accuracy; these values were 57% and 78%, respectively, for cholangioscopic-directed biopsies. Each procedure had 100% specificity. CONCLUSIONS: Analysis of bile duct biopsies is important in management of patients with indeterminate biliary strictures. Use of a special handling protocol for these small biopsies could reduce the number of cases with insufficient material for diagnosis. Increasing the sample size (either by using larger biopsy forceps or obtaining more biopsy bites) could improve the sensitivity of the SpyGlass technique. As endoscopists and pathologists gain more experience in collecting and handling small biopsies, the diagnostic efficacy of intraductal biopsies will continue to improve.

Non-hematopoietic allograft cells directly activate CD8+ T cells and trigger acute rejection: an alternative mechanism of allorecognition.

Despite evidence that human non-hematopoietic cells, such as vascular endothelium, can activate allogeneic T lymphocytes in vitro, the prevailing view has been that hematopoietic antigen-presenting cells are required to trigger alloimmune responses in vivo. Here we report that mouse non-hematopoietic cells activate alloreactive CD8+ T lymphocytes in vitro and in vivo. We also show that vascularized cardiac allografts are acutely rejected via CD8+ direct allorecognition even if the alloantigen is not presented by hematopoietic professional antigen-presenting cells. Because activation of alloreactive CD8+ T cells by donor-type non-hematopoietic cells can continue for the life of the allograft, these findings present a new clinically relevant mechanism of allorecognition and should be taken into consideration when developing strategies to prevent allograft vasculopathy or to induce tolerance.